The present invention, in some embodiments thereof, relates to methods of culturing and expanding mesenchymal stem cells and isolated cell populations generated thereby.
Mesenchymal stem cells (MSCs) are non-hematopoietic cells that are capable of differentiating into specific types of mesenchymal or connective tissues including adipose, osseous, cartilaginous, elastic, neuronal, hepatic, pancreatic, muscular, and fibrous connective tissues. The specific differentiation pathway which these cells enter depends upon various influences from mechanical influences and/or endogenous bioactive factors, such as growth factors, cytokines, and/or local microenvironmental conditions established by host tissues.
MSCs reside in a diverse host of tissues throughout the adult organism and possess the ability to ‘regenerate’ cell types specific for these tissues. Examples of these tissues include adipose tissue, umbilical cord blood, periosteum, synovial membrane, muscle, dermis, pericytes, blood, bone marrow and trabecular bone.
The multipotent character of mesenchymal stem cells make these cells an attractive therapeutic tool and candidate for transplantation, capable of playing a role in a wide range of clinical applications in the context of both cell and gene therapy strategies. Mesenchymal cells may be used to enhance hematopoietic engraftment post-transplantation, to correct inherited and acquired disorders of bone and cartilage, for implantation of prosthetic devices in connective and skeletal tissue, and as vehicles for gene therapy.
Even though MSCs multiply relatively easily in vitro, their proliferative potential and their stem cell characteristics are continuously decreased during prolonged culture. For example, it has been shown that expansion in culture leads to premature senescence (the process of aging characterized by continuous morphological and functional changes). Cells became much larger with irregular and flat shape and the cytoplasm became more granular. These senescence-associated effects are continuously acquired from the onset of in vitro culture (PLoS ONE, May 2008|Volume 3|Issue 5|e2213). As a result, the successful manufacturing for commercialization of large batches from one donor of homogenous MSCs that preserve their characteristics following expansion in culture remains a challenge.
Due to the low or absent expression of MHC molecules, especially class II on mesenchymal stem cells, these cells may be considered immune privileged, thus paving the way for allogeneic transplantation of such cells for the treatment of a wide range of disorders. Accordingly, improved methods of expanding banks of mesenchymal stem cells have become an important factor for commercializing their use.
StemRegenin 1 (SR1), a purine derivative, has been identified in an unbiased screen to promote the ex vivo expansion of CD34+ cells. Mechanistic studies show that SR1 acts by antagonizing the aryl hydrocarbon receptor (AhR) expansion [Boitano et al. 2010 Science 329(5997):1345-1348].
U.S. Patent Application No. 20100183564 discloses the use of aryl hydrocarbon receptor antagonists for hematopoietic stem cell expansion. U.S. Patent Application No. 20100183564 does not suggest or mention the use of aryl hydrocarbon receptor antagonists for non-hemaptopoietic stem cell expansion.
Additional background art includes WO 03/062369, U.S. Patent Application No. 20050260748 and Farre et al., Growth Factors, 2007 April; 25(2):71-6.